Thymidylate kinase (TMPK) is a key enzyme for the phosphorylation of dTMP to dTDP, which is converted to dTTP by nucleotide diphosphate kinases (NDPKs) for DNA synthesis. Ribonucleotide reductase (RNR) is the enzyme essential for de novo synthesis of dNTPs. RNR-catalyzed reactions generate dADP, dGDP, dCDP and dUDP from the corresponding NDPs (Nordlund and Reichard, 2006). Therefore, dTDP is the only dNDP not directly derived from RNR reaction, but from TMPK reaction.
Conventional anti-cancer therapies often directly induce genotoxicity (Garg et al., 2010). For example, thymidylate synthase (TS) inhibitor, 5-FU or 5-FdUrd, blocks the conversion of dUMP to dTMP, causing dUTP to accumulate and 5-FdUTP formation (Longley et al., 2003). Since DNA polymerases cannot discriminate between dUTP and dTTP (Bessman et al., 1958; Mosbaugh, 1988), excessive amounts of dUTP and 5FdUTP are mis-incorporated into DNA, triggering genotoxicity-induced cell death (Ahmad et al., 1998). Consequently, such anti-metabolites produce excessive DNA damage due to erroneous nucleotide incorporation and causes cancer cells death while being highly toxic to normal cycling cells (Ahmad et al., 1998). In contrast, blocking TMPK does not cause dUTP accumulation, therefore the general cytotoxicity is much lower than TS inhibition (Hu et al, 2012)
It is known that double-strand breaks (DSBs) in proliferating cells are mainly repaired by homologous recombination (HR) in which the repair of a single DSB needs more than 10 thousands of dNTPs new incorporation (Robert et al., 2011; San Filippo et al., 2008). As such, RNR function in supply of dNTPs is critical for HR repair (Burkhalter et al., 2009). Of note, blocking RNR on its own induces DNA damage signal and replication stress (Helleday et al., 2008). Since dTDP formation specifically requires TMPK function, we showed that blocking TMPK decreases the efficiency of DSBs repair and sensitizes tumor cells to genotoxic insults (Hu et al., 2008, 2012)
Accordingly, inhibition of TMPK that blocks dTDP conversion from dTMP will inhibit DNA replication and repair that demand a large quantity of dTTP. Therefore, the TMPK inhibitor can be used for chemosensitization and inhibiting the growth of cancer. The TMPK inhibitor treatment has low general genotoxicity as compared to 5 FU or blocking dTMP formation.
The main problem with chemotherapy is the lack of differentiation between tumor- and rapidly-dividing cells in normal tissues. This causes substantial side effects, which can in the long run lead to secondary cancers induced by the treatment. Chemotherapeutic agents often cause unwanted general cytotoxicity, and activities of several DNA repair pathways enable tumor cells to survive by removing lesions (Helleday et al., 2008).
Small-molecule inhibitors of checkpoint pathways or DNA repair machineries were identified and used as cellular radio- and chemosensitization compounds in clinical trials (Bolderson et al., 2009). Given the differences in checkpoint and DNA repair alteration during tumorigenesis, the therapeutic efficacies of these strategies were found various depending on the checkpoint context of tumor (Jackson and Bartek, 2009; Jiang et al., 2009).
Moreover, derangement of DNA damage response could cause the accumulation of DNA error during therapy, which might provoke secondary tumor development (Mimeault et al., 2008). Therefore, it is important to develop a chemosensitization regimen that does not disrupt the checkpoint network while specifically inducing cancer cell death with little side effect.